This project concerns mechanisms of cross-linked envelop formation in epidermal cells (keratinocytes). Serially cultivated with support from feeder layers of lethally irradiated mouse 3T3 cells (the Rheinwald-Green system), the keratinocytes retain differentiated function and appear analogous to those of normal stratified squamous epithelia. The overall objectives of this research are two-fold: 1. to characterize and elucidate interactions among the three components participating in envelope formation (envelope subunit protein, plasma membrane where the subunit protein localizes, and transglutaminase), and 2. to explore states of cellular physiology in which these interactions are altered or abnormal. To identify elements of plasma membrane (lipid, protein) responsible for envelope protein localizations, efforts are under way to study binding of the protein to membranes. Separation of transglutaminase into purified isozymes may help determine whether a specific form is involved in envelop cross-linking. Keratinocyte lines from human epidermal and oral squamous cell carcinomas are being examined for variants that exhibit defective envelope formation. A line that exhibits envelope precursor protein but improper localization or lacks one transglutaminase isozyme is of great potential usefulness. Chemical agents that alter cell differentiation or growth will also be explored for effects on envelope assembly.